The quantitative determination of analytes in body fluids is of great importance in the diagnoses and maintenance of certain physiological abnormalities. Some non-limiting examples of analytes that may be monitored in certain individuals include cholesterol (e.g., HDL, LDL, total), triglycerides, globulin, albumin, total protein, blood urea nitrogen, creatinine, and alkaline phosphastase.
Nuclear magnetic resonance (NMR) spectroscopy is an analytical and diagnostic technique that is used for structural and quantitative analysis of one or more analytes in a sample. One specific type of NMR is time domain nuclear magnetic resonance (TD-NMR). TD-NMR spectrometers are used for spin-lattice (T1) and spin-spin (T2) relaxation measurements. NMR spectrometers have been used to analyze body fluid samples, such as blood plasma. NMR is based on nuclear magnetic properties of certain elements and isotopes of those elements. One such element that is commonly analyzed by NMR is hydrogen, which has a single proton and an intrinsic nuclear spin. Hydrogen is present in many analytes of interest and has high natural abundance. When hydrogen nuclei are placed in a magnetic field, they adopt one of two allowed orientations. Therefore, the resulting magnetic moment can be aligned with the magnetic field or opposed to the magnetic field. The two orientations are separated by an amount of energy that depends on the strength of the magnetic field and the strength of the interaction between the hydrogen nucleus and the field. The energy difference may be determined by applying an electromagnetic pulse at a characteristic resonance frequency, which causes the nuclei aligned with the field (lower energy state) to align against the field (higher energy state).
The resonance frequency, ν, of a hydrogen nucleus is dependent on the strength of the magnetic field, Bo, and is given by:ν=γBo/2πwhere ν is in units of MHz, Bo is in units of tesla (T) and γ is the fundamental gyromagnetic ratio for hydrogen (1H) and is equal to 267.512×106 rad T−1s−1. Magnetic field strengths commonly used for NMR spectroscopy are in the range of from 1.4 to 14.1 T, corresponding to hydrogen resonance frequencies of from 60 to 600 MHz. Since hydrogen is the most common nucleus studied, NMR spectrometers are often classified by their hydrogen resonance frequencies instead of their actual magnetic field strengths.
The two most common types of magnets used in NMR spectrometers are permanent magnets and superconducting magnets. Although permanent magnets provide acceptable field stability and are less costly, the field strength is limited to approximately 1.4 T (60 MHz).
In contrast, superconducting magnets can provide much higher fields in the range of from about 4.7 to about 18.8 T (from 200 to 800 MHz). It is important to remember that the resonance frequencies of hydrogen nuclei, as well as other non-identical nuclei, are proportional to the field strength. Neighboring nuclei, in the same molecule or in the solvent, may greatly impact the resonance frequency of a particular hydrogen nucleus. Therefore, to obtain characteristic NMR spectra with high resolution, it is desirable to use the higher magnetic fields achieved with superconducting magnets. Unfortunately, NMR systems that utilize superconducting magnets are very expensive, require cryogenic cooling with liquid nitrogen and liquid helium, and are very large (commonly occupying an entire room). In addition to field strength, the stability and homogeneity of the magnetic field should be controlled to obtain high-quality NMR spectra. High-field NMR spectrometers also employ special locking electronics to compensate for small field instabilities. In the sample probe, additional electronic hardware is used to control the homogeneity of the magnetic field by a process called shimming.
Additionally, the resulting high-resolution NMR spectra are complex and must be interpreted by a highly-trained scientist. The individuals who operate the high-resolution NMR equipment need to be well-trained. High-resolution NMR spectroscopy typically involves acquiring a complete spectrum and identifying peaks for qualitative and quantitative analysis.
Therefore, it would be desirable to provide a method of determining the concentration of one or more analytes using NMR that provides lower costs and is a convenient method to use without requiring highly-trained operators or scientists.